Structural analysis of MDM2 RING separates degradation from regulation of p53 transcription activity

Nomura, Koji; Klejnot, Marta; Kowalczyk, Dominika; Höck, A.; Sibbet, Gary; Vousden, Karen H.; Huang, Danny T.

doi:10.1038/nsmb.3414

Cited by 51 publications

(102 citation statements)

References 63 publications

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“…These residues are mainly located at the surface of hMEX‐3C Ring finger domain and may be responsible for the recruitment of E2–Ub. The Residues I440 and R479 in MDM2 play key role to interact with E2 and R479 facilitates to stabilize the closed active E2–Ub conformation . Hence, we identified that I610 and Q648 in hMEX‐3C may play key role to interact with E2 and stabilize ubiquitin in the closed active E2–Ub conformation.…”

Section: Resultsmentioning

confidence: 79%

“…Given that the overall structure of hMEX‐3C Ring finger domain closely resembles the Ring finger domain of MDM2, we speculated that the Ring finger domain of hMEX‐3C may interact with UbcH5b via the same binding site MDM2 used. The interaction between MDM2 Ring finger domain and UbcH5b–Ub showed that several key residues of MDM2/MDMX play a crucial role in stabilizing the closed UbcH5b–Ub conformation . According to the sequence alignment [Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Interestingly, mutations of V609 and V646 to glutamic acid totally abolish the ubiquitination activity of hMEX‐3C Ring finger domain [Fig. (d)], which indicates the requirement of extended nature of the surface in regulating the functional interaction with the E2 . The ubiquitination assays of hMEX‐3C revealed that the entire Ring finger domain, together with specific E2s, plays a remarkable role in its self‐ubiquitination [Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Recent studies have reported that the Ring finger domain binds E2–Ub thioester intermediate and promotes E2–Ub closed active conformations required for direct Ub transfer from E2 to substrate . The interaction between E2–Ub and the Ring finger domain has also been investigated by structural research . Previous structural studies have reported that the Ring finger domain interacts to the E2 and positions the “donor ubiquitin” in the E2–Ub for ubiquitin transfer to a remotely bound protein substrate .…”

Section: Introductionmentioning

confidence: 99%

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Structural and functional characterization of hMEX‐3C Ring finger domain as an E3 ubiquitin ligase

et al. 2018

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MEX‐3C, a novel RNA binding E3 ubiquitin ligases, contains two N‐terminal heterogeneous nuclear ribonucleoprotein K homology (KH) domains and C‐terminal Ring finger domain. Recent evidence has suggested that human MEX‐3C has a strong bondage with carcinogenesis and the MEX‐3C‐mediated ubiquitination of RIG‐I is essential for the antiviral innate immune response. Moreover, the Ring finger domain of MEX‐3C could regulate the degradation of HLA‐A2 (an MHC‐I allotype) mRNA with a novel mechanism. However, the structural basis for the ubiquitination catalyzed by hMEX‐3C Ring finger domain remains evasive. In this study, we solved the crystal structure of dimeric Ring finger domain of hMEX‐3C and compared it with the complex structure of MDM2/MDMX–UbcH5b–Ub. Our ubiquitination assay demonstrated that the Ring finger domain of hMEX‐3C acts as a ubiquitin E3 ligase in vitro, cooperating with specific E2 to mediate ubiquitination. Then, we identified several key residues in Ring finger domain of hMEX‐3C possibly involved in the interaction with E2–Ub conjugate and analyzed the E3 ligase activities of wild type and mutants at key sites. Additionally, zinc chelation experiments indicated that the intact structural stability is essential for the self‐ubiquitination activity of the Ring finger domain of hMEX‐3C. Taken together, our studies provided new insight into the mechanism of the Ring finger domain of hMEX‐3C that may play an important role in eliciting antiviral immune responses and therapeutic interventions.

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Section: Resultsmentioning

confidence: 79%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Structural and functional characterization of hMEX‐3C Ring finger domain as an E3 ubiquitin ligase

et al. 2018

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“…(a) Ribbon representation of a single RNF4 monomer from UBE2D1~Ub:RNF4 complex (PDB: 4AP4) showing the zinc ions as spheres, the residues involved in zinc‐coordination and the allosteric linchpin residue as ball‐and‐sticks, and demarcating the E2‐binding site. The ribbon diagram is colored to reflect the average pairwise positional shift of each overlapping C α atom between RNF4 and unique RING cores from all available E2–RING E3 complex structures including RNF146 (PDB: 4QPL), TRIM25 (PDB: 5FER), BIRC2 (PDB: 6HPR), BIRC3 (PDB: 3 EB6), BIRC7 (PDB: 4AUQ), RNF38 (PDB: 4V3K), RNF25 (PDB: 5D1M), RNF165 (PDB: 5D0M), MDM2 (PDB: 5MNJ), TRIM23 (PDB: 5VZW), RNF13 (PDB: 5ZBU), E4B (PDB: 3L1Z), RNF2 (PDB: 3RPG), GP78 (PDB: 2LXP), SIZ1 (PDB: 5JNE), c‐CBL (PDB: 1FBV), RBX1 (PDB: 4P5O), TRAF6 (PDB: 3HCT), TRIM5 (PDB: 4TKP), RNF8 (PDB: 4WHV), ZNRF1 (PDB: 5YWR), LNX1 (PDB: 5H7S), CHIP (PDB: 2C2V), and FANCL (PDB: 2CCG) . (b) UBE2D1~Ub:RNF4 complex highlighting position of the RING domain relative to the E2, with the allosteric linchpin residue coordinating E2 as well as the donor Ub shown as ball‐and‐sticks.…”

Section: Ring E3 Morphologymentioning

confidence: 99%

Structural basis of generic versus specific E2–RING E3 interactions in protein ubiquitination

Gundogdu

Walden

2019

Protein Science

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Protein ubiquitination is a fundamental regulatory component in eukaryotic cell biology, where a cascade of ubiquitin activating (E1), conjugating (E2), and ligating (E3) enzymes assemble distinct ubiquitin signals on target proteins. E2s specify the type of ubiquitin signal generated, while E3s associate with the E2~Ub conjugate and select the substrate for ubiquitination. Thus, producing the right ubiquitin signal on the right target requires the right E2–E3 pair. The question of how over 600 E3s evolved to discriminate between 38 structurally related E2s has therefore been an area of intensive research, and with over 50 E2–E3 complex structures generated to date, the answer is beginning to emerge. The following review discusses the structural basis of generic E2–RING E3 interactions, contrasted with emerging themes that reveal how specificity can be achieved.

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MANF attenuates neuronal apoptosis and promotes behavioral recovery via Akt/MDM‐2/p53 pathway after traumatic spinal cord injury in rats

Gao

Fan³

et al. 2018

BioFactors

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The aim of this study was to investigate the potential effect and mechanism of action of MANF in attenuating neuronal apoptosis following t-SCI. A clip compressive model was used to induce a crush injury of the spinal cord in a total of 230 rats. The Basso, Beattie, and Bresnahan (BBB) score, spinal cord water content, and blood spinal cord barrier (BSCB) permeability were evaluated. The expression levels of MANF and its downstream proteins were examined by western blotting. Immunofluorescence staining of MANF, NeuN, GFAP, Iba-1, cleaved caspase-3, and TUNEL staining were also performed. Cells were counted in six randomly selected fields in the gray matter regions of the sections from two spinal cord sites (2 mm rostral and caudal to the epicenter of the injury) per sample. A cell-based mechanical injury model was also conducted using SH-SY5Y cells. Cell apoptosis and viability were assessed by flow cytometry, an MTT assay, and trypan blue staining. Subcellular structures were observed by transmission electron microscopy. MANF was mainly expressed in neurons. The expression levels of MANF, and its downstream target, p-Akt, were gradually increased and after t-SCI. Treatment with MANF increased Bcl-2 and decreased Bax and CC-3 levels; these effects were reversed on treatment with MK2206. The BBB score, spinal cord water content, and BSCB destruction were also ameliorated by MANF treatment. MANF decreases neuronal apoptosis and improves neurological function through Akt/MDM-2/p53 pathway after t-SCI. Therefore, MANF might be a potential treatment for patients with t-SCI.© 2018 BioFactors, 2018.

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Structural analysis of MDM2 RING separates degradation from regulation of p53 transcription activity

Cited by 51 publications

References 63 publications

Structural and functional characterization of hMEX‐3C Ring finger domain as an E3 ubiquitin ligase

Structural and functional characterization of hMEX‐3C Ring finger domain as an E3 ubiquitin ligase

Structural basis of generic versus specific E2–RING E3 interactions in protein ubiquitination

MANF attenuates neuronal apoptosis and promotes behavioral recovery via Akt/MDM‐2/p53 pathway after traumatic spinal cord injury in rats

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